The long-term objective of this application is to deliver a unique biomaterial that can easily be molded into place by a surgeon, and will resorb as it induces rapid tissue regeneration. Toward this objective, we invented a biomaterial based on colloidal gel technology, which we have demonstrated to be effective in calvarial defect regeneration. The key feature that distinguishes colloidal gels from the two major classes of scaffolding biomaterials (hydrogels and solid scaffolds) is their paste-like rheology, which in turn is attributed to electrostatic interaction of the nanoparticle constituents. Although this new class of scaffolds is highly versatile in its unbounded combination of possible synthetic and natural nanoparticles, we have elected to focus on a combination of naturally occurring materials with controlled release of bioactive signals. Therefore, the objective of this project is to develop a malleable material that can be spread into place in a cranial defect, while releasing bioactive factors and allowing native bone to penetrate and resorb the material. The corresponding central hypothesis is that the growth factor-loaded colloidal gels will regenerate bone in cranial defects significantly faster and more completely than unloaded colloidal gels or commercial hydroxyapatite bone fillers. To test this hypothesis, we propose three specific aims: 1) to synthesize and characterize novel colloidal gels with modulated rheological properties, 2) to engineer and refine colloidal gels in vitro, and 3) to determine the efficacy of colloidal gels in a rat cranial defect model. Building on our published characterization of prototype colloidal gels, our overall strategy will be to significantly expand our repertoire first by evaluating the rheological and release properties of a variety of combinations of specific sulfated glycosaminoglycans (GAGs, negatively charged) and hydroxyapatite nanoparticles (positively charged), which have been identified as an internally cohesive colloidal gel network. A specific subset of these combinations will then be thoroughly evaluated in vitro for their efficacy in promoting osteogenesis with rat bone marrow-derived mesenchymal stem cells (BMSCs). The most promising groups from these in vitro studies will be evaluated in critical-sized rat calvarial defects, with the project thereby culminating in the identification of the leading combination of GAGs, hydroxyapatite, and osteogenic and angiogenic signals for calvarial defect regeneration. Successful completion of this project will lay the foundation for an entirely new sub-field for tissue engineering scaffolding biomaterials. The true impact of this line of research is its extraordinary versatility and relatively straightforward set of design principles as a means to create bioresorbable, pastes of tunable consistency, with the capability for controlled release of bioactive signals. We and other investigators world- wide will be able to explore a seemingly infinite number of innovative combinations of interactive nanoparticles for applications beyond calvarial defect regeneration, from osteochondral regeneration to liver regeneration to any other conceivable application where such a material is desired.